补体C3a受体在盲肠结扎穿孔致脓毒症大鼠认知障碍中的作用及机制

doi:10.16098/j.issn.0529-1356.2023.06.007

摘要/Abstract

摘要：

目的探讨补体C3a受体（C3aR）在盲肠结扎穿孔（CLP）致脓毒症大鼠认知障碍中的作用及潜在的作用机制。方法 132只大鼠随机分为假手术组（sham组）和脓毒症组（CLP组），每组的初始数量为66只动物，每个时间点设置22只动物。构建SD大鼠CLP动物模型，于术后1、15和30 d取血清和脑（海马和前额叶皮质），采用ELISA法测定肿瘤坏死因子α（TNF-α）、白细胞介素1β（IL-1β）和IL-6水平。Western blotting检测脑标本中Tau蛋白（Tau）、磷酸化Tau（p-Tau）和C3aR蛋白表达。将66只大鼠随机分为3组：sham组、CLP组和C3aR拮抗剂(C3aRa)干预组。于CLP后第15、17和19天，C3aRa干预大鼠于CLP术后第30天接受抑制性回避测试和物体识别测试，并采用免疫荧光法检测海马中C3aR、离子钙接头蛋白分子1（Iba1）、胶质纤维酸性蛋白（GFAP）和p-Tau表达。结果与sham组相比，CLP组大鼠血清及脑组织中TNFα、IL-1β和IL-6在CLP后30 d内先增加后减少。在CLP组海马组织和前额叶皮层中分别观察到术后30 d和术后1、15 d Tau磷酸化显著增强（P<0.05）。与sham组相比，CLP组术后15 d和30 d在大鼠海马和前额叶皮层中均观察到C3aR蛋白水平显著增加（P<0.05）。与CLP组相比，C3aRa干预后大鼠海马内源性C3aR含量明显降低（P<0.05），Iba1、GFAP和p-Tau免疫染色明显抑制（P<0.05）。C3aRa干预CLP大鼠可逆转认知功能损伤。结论 C3aR在脓毒症诱导神经退行性过程中相关生物化学和行为学变化发展中发挥重要作用。通过在海马注射C3aRa能够改善由脓毒症引发的神经退化过程。

关键词: 补体C3a受体, 脓毒症, 认知障碍, 炎症, 免疫印迹法, 大鼠

Abstract:

Objective To study the role of complement C3a receptor (C3aR) in cognitive impairment in rats with sepsis induced by cecal ligation and puncture (CLP), and to explore the possible mechanism. Methods Totally 132 rats were randomly divided into sham operation group (sham group) and sepsis group (CLP group). The initial number of each group was 66 animals, and 22 animals were set at each time point. The SD rat CLP animal model was constructed, and serum and brain (hippocampus and prefrontal cortex) samples were collected at day 1, day 15 and day 30 after operation. The levels of tumor necrosis factor-α(TNF-α), interleukin-1β (IL-1β) and IL-6 in the samples were determined by ELISA. Western blotting detected Tau protein (Tau), phosphorylated Tau（p-Tau）and C3aR expression in brain samples. Totally 66 rats were randomly divided into 3 groups, sham operation group, CLP group and C3aR antagonist(C3aRa) intervention group. On 15th, 17th, and 19th days after CLP, C3aRa intervened in rats, and they received inhibition avoidance test and object recognition test 30 days after CLP. The expressions of C3aR, lonized calcium binding adapter molecule 1(Iba1), GFAP and p-Tau in the hippocampus were detected by immunofluorescence. Results Compared with the sham group, the serum and brain tissue (TNFα, IL-1β and IL-6) of rats in the CLP group first increased and then decreased within 30 days after CLP. In the hippocampus and prefrontal cortex of CLP group, Tau phosphorylation was significantly enhanced at day 30 and day 1 and 15 after surgery, respectively (P<0.05). Compared with the sham group, C3aR protein levels in hippocampus and prefrontal cortex of rats in CLP group increased significantly at day 15 and 30 after operation (P<0.05). Compared with the CLP group, the endogenous C3aR content decreased significantly after C3aRa intervention (P<0.05), and Iba1, GFAP and p-Tau immunostaining were significantly inhibited (P<0.05). The C3aRa intervention in CLP rats reversed the cognitive impairment. Conclusion C3aR plays an important role in the development of biochemical and behavioral changes commonly associated with the onset of sepsis-induced neurodegenerative processes. C3aRa can be injected into the hippocampus to counteract the neurodegenerative process induced by sepsis.

Key words: Complement C3a Receptor, Sepsis, Cognitive impairment, Inflammation, Western blotting, Rat

中图分类号:

R714

徐颖刘斌郑兴何宗钊.

补体C3a受体在盲肠结扎穿孔致脓毒症大鼠认知障碍中的作用及机制 [J]. 解剖学报, 2023, 54(6): 668-675.

XU Ying LIU Bin ZHENG Xing HE Zong-zhao. Role and mechanism of complement C3a receptor in the cognitive impairment of septic rats induced by cecal ligation and perforation[J]. Acta Anatomica Sinica, 2023, 54(6): 668-675.

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